Alzheimer's disease (AD) is a neurodegenerative disease and the most common cause of dementia. Early memory problems and gradual and progressive decline in cognitive functions beyond normal ageing are characteristic for AD. Post-mortem studies have shown the neuropathological hallmarks of the disease include extracellular amyloid plaques mainly consisting of 38 to 43 amino acids long peptides called AP peptide and intracellular neurofibrillary tangles with hyperphosphorylated TAU protein as the characteristic component.
Aβ peptides are generated in the amyloidogenic pathway from the Amyloid Precursor Protein (APP). In this pathway, Aβ peptides are generated by the sequential action of two proteases, β- and γ-secretase. The β-secretase activity is exerted by the (3-site APP cleaving enzyme 1 (BACE1) and BACE1 mediated APP cleavage results in shedding of the extracellular APP ectodomain (sAPPβ). The remaining membrane-bound C-terminal fragment (C99) is further processed by γ-secretase, which catalyzes an unusual proteolysis within the transmembrane region, resulting in the release of the APP intracellular domain (AICD) in the cytosol and the exocytosis of Aβ peptides in the extracellular environment. The majority of Aβ produced is 40-amino acid residues in length (Aβ40). Although the 42-residue form (Aβ42) is a minor species, it more readily aggregates to produce fibrils and ultimately amyloid plaques.
Next to the pathology also human genetics studies strongly suggest that Aβ plays a central role in AD pathogenesis. Today, over 200 autosomal dominant mutations that cause familial AD (FAD) have been found in the genes for APP and presenilin, the active subunit of γ-secretase. These mutations invariably lead to either increased Aβ42 to Aβ40 ratio or over-production of total Aβ. Notably, the FAD mutations in APP are found near the β- and γ-secretase cleavage sites and make APP a more efficient substrate for endoproteolysis by the secretases. Of particular relevance here are the K670N; M671L (Swedish) double mutation and the A673V mutation that are adjacent to the β-secretase cleavage site and cause FAD by increasing β-secretase processing and total Aβ production. Interestingly genetic variants have been identified that protect against AD. A low-frequency mutation in APP, the A673T coding substitution, was recently shown to be associated with decreased risk of AD and reduced cognitive decline in the elderly (Jonsson et al. 2012, Nature 488, 96-99). APP harboring the A673T substitution—located two amino acids C-terminal to the β-secretase cleavage site is less efficiently cleaved by β-secretase, leading to a ˜40% reduction in Aβ production in vitro.
Cleavage of APP by Beta-site APP Cleaving Enzyme1 (BACE1) is the rate limiting step in the generation of the Aβ peptide. BACE1 is a membrane-bound aspartyl protease that is optimally active at a slightly acidic pH. Although BACE1 is localized in various organelles, its activity is reported to be at a maximum in endosomes and to a lower extend in the trans golgi network (TGN), hence most APP is cleaved by BACE1 in the endocytic compartment. Evidence that BACE-1 is the sole β-secretase activity in the brain was provided by the observations that BACE-1 knockout mice completely lacked both β-secretase enzyme activity and the product of β-cleavage, CTF99 (Roberds et al., 2001, Hum.
Mol. Genet. 10, 1317-1324, Luo et al., 2001, Nat. Neurosci. 4, 231-232). Ongoing clinical trials with BACE1 inhibitors confirm that BACE1 is the sole β-secretase activity in human brain, since pharmacological BACE1 inhibition blocks Aβ production.
Soon after the discovery of BACE1, a related membrane-bound aspartic protease BACE2 was identified that shares 64% amino acid similarity to BACE1. Although BACE2 can generate Aβ in vitro, it appears not to do so in vivo as mentioned above. BACE1 and its homologue BACE2 are members of the pepsin-like family of aspartic proteases (cathepsin D and E, pepsin A and C, renin, napsin A). They display a typical bilobal structure with the catalytic site located at the interface between the N- and the C-terminal lobe (Hong et al, 2000, Science 290, 150-153, Ostermann et al, 2006, Journal of molecular biology, 355, (2), 249-61). BACE1 and 2 are anchored to the cell membrane via a transmembrane domain, which, together with several unique amino acid stretches and the arrangement of the three disulfide bridges (Haniu et al., 2000, J. Biol. Chem. 275, 21099-21106) sets BACE apart from the rest of the pepsin family and facilitates the generation of relatively specific inhibitors for BACE1 and 2.
Next to APP a variety of CNS and peripheral BACE1 substrates and associated functions have been described (Hemming et al. 2009, PLoS ONE 4, e8477, Kuhn et al. 2012, EMBO J. 31, 3157-3168; Zhou et al. 2012, J. Biol. Chem. 287, 25927-25940, Stutzer et al. 2013, J. Biol. Chem. 288, 10536-10547, reviewed in Vassar et al., J. Neurochem. (2014) 10.1111/jnc.12715). Examples of BACE1 substrates are L1, CHL1, GLG1, PAM, SEZ6, SEZ6L, Jag1, NRG1, NaVβ2, VEGFR1 and APLP1. Consequently BACE1 has a wide variety of potential physiologic functions including, but not exclusively in cell differentiation, immunoregulation, myelination, synaptic development and plasticity, cell death, neurogenesis and axonal guidance (Wang et al. Trends in Pharmacological Sciences, April 2013, Vol 34, No. 4, pp. 215-225; Yan and Vassar Lancet Neurol. 2014, Vol. 13, pp. 319-329; Yan et al. J Alzheimers Dis. 2014, Vol. 38, No. 4, pp. 705-718).
For example in BACE1 knock-out mice, loss of cleavage of neuregulin 1 (NRG1) type III resulted in impaired post-natal myelination in the PNA and CNS (Fleck et al. 2012, Curr. Alzheimer Res. 9, 178-183; Willem et al. 2009, Semin. Cell Dev. Biol. 20, 175-182). Loss of cleavage of NRG1 type I results in abnormal muscle spindle formation and maintenance and associated defects in coordinated movement (Cheret et al. 2013). BACE1 processing of β-subunits of voltage-gated sodium channels controls cell-surface NaV channel density, neuronal excitability, and seizure susceptibility (Kim et al. 2011, J. Biol. Chem. 286, 8106-8116). BACE1-dependent CHL1 cleavage is known to be involved in axon outgrowth and neuronal survival (Naus et al. 2004, J. Biol. Chem. 279, 16083-16090). BACE1-dependent Jag1 cleavage regulates post-natal neurogenesis and astrogenesis by modulating Notch 1 signalling.
Therefore, in addition to Alzheimer's disease, Down syndrome, and related diseases, BACE inhibition may find therapeutic and/or prophylactic treatment use in conditions such as traumatic brain injury (TBI), temporal lobe epilepsy (TLE), hypoxia, ischemia, cellular stress, neuroinflammatory disorders, disruptions in cerebral metabolism, age-related macular degeneration, Sjogren syndrome, Spinocerebellar ataxia 1, Spinocerebellar ataxia 7, Whippel's disease and Wilson's disease, age-related macular degeneration, amyotrophic lateral sclerosis (ALS), arterial thrombosis, autoimmune/inflammatory diseases, cardiovascular diseases such as myocardial infarction and stroke, dermatomyositis, gastrointestinal diseases, Glioblastoma multiforme, Graves' Disease, Huntington's Disease, inclusion body myositis (IBM), inflammatory reactions, Kaposi Sarcoma, Kostmann Disease, lupus erythematosus, macrophagic myofasciitis, juvenile idiopathic arthritis, granulomatous arthritis, malignant melanoma, multiple myeloma, and rheumatoid arthritis.
Also BACE2 has a broad expression profile, with relative high expression levels in most different cell types and organs in the periphery and lower level of expression in astrocytes in the brain. As mentioned above also BACE2 has a broad spectrum of substrates as exemplified by the study in the pancreatic islets mentioned above (Stutzer et al. 2013).
BACE2 is expressed in pancreatic 13 cells, where it cleaves Tmem27 (Esterházy et al. Cell Metabolism 2011). Inhibition of BACE2 therefore may provide a potential mechanism to result in increased β cell mass, and a potential mode of action in the treatment or prevention of Type2 diabetes
BACE2 is also known to be involved in the cleavage of APP (Wang et al. Trends in Pharmacological Sciences, April 2013, Vol. 34, No. 4, pp. 215-225), IL-1R2 (Kuhn et al. J. Biol. Chem. 2007, Vol. 282, No. 16, pp. 11982-11995), and pigment cell-specific melanocyte protein (PMEL) (Rochin et al. PNAS, Jun. 25, 2013, Vol. 110, No. 26, pp. 10658-10663), therefore indicating a potential application for BACE2 inhibitors in the treatment of Down's syndrome, hypertension, malignant melanoma and multiple melanoma. BACE2 is unregulated in human breast cancers (Kondoh et al. Breast Cancer Res. Treat., 2003, Vol. 78, pp. 37-44), and therefore BACE2 inhibitors may provide a potential in the treatment of breast cancers.
Inhibitors of BACE1 and/or BACE2 may thus be useful for the therapeutic and/or prophylactic treatment of Alzheimer's disease (AD), mild cognitive impairment, senility, dementia, dementia with Lewy bodies, Down's syndrome, dementia associated with stroke, dementia associated with Parkinson's disease, dementia of the Alzheimer's type, dementia associated with beta-amyloid. In addition to Alzheimer's disease, and related diseases, BACE inhibition may find therapeutic and/or prophylactic treatment use in conditions such as traumatic brain injury (TBI), temporal lobe epilepsy (TLE), hypoxia, ischemia, cellular stress, neuroinflammatory disorders, disruptions in cerebral metabolism, age-related macular degeneration, Sjogren syndrome, Spinocerebellar ataxia 1, Spinocerebellar ataxia 7, Whippel's disease and Wilson's disease, age-related macular degeneration, amyotrophic lateral sclerosis (ALS), arterial thrombosis, autoimmune/inflammatory diseases, cardiovascular diseases such as myocardial, infarction and stroke, dermatomyositis, gastrointestinal diseases, Glioblastoma multiforme, Graves' Disease, Huntington's Disease, inclusion body myositis (IBM), inflammatory reactions, Kaposi Sarcoma, Kostmann Disease, lupus erythematosus, macrophagic myofasciitis, juvenile idiopathic arthritis, granulomatous arthritis, malignant melanoma, multiple myeloma, and rheumatoid arthritis, type 2 diabetes and other metabolic disorders, hypertension, malignant melanoma and multiple melanoma and breast cancer.
WO 2012/117027 (Janssen Pharmaceutica NV) discloses 4-amino-6-phenyl-6,7-dihydropyrazolo[1,5-a]pyrazine derivatives as BACE inhibitors. WO 2012/057247 (Shionogi & Co., Ltd.) describes fused aminodihydropyrimidine derivatives useful as BACE inhibitors.